Deng Yunlan, Wu Fengli, Li Qianqian, Yao Lidie, Yang Chengzhi, Ma Long, Yao Xinsheng, Li Jun
Department of Immunology, Center of Immunomolecular Engineering, Innovation & Practice Base for Graduate Students Education, Zunyi Medical University, Zunyi, China.
Department of Laboratory, The Affiliated Yongchuan Hospital of Chongqing Medical University, Chongqing, China.
Front Immunol. 2025 Jan 20;15:1503788. doi: 10.3389/fimmu.2024.1503788. eCollection 2024.
Immunoglobulin (Ig) refers to the nomenclature for all antibody proteins produced by B lymphocytes. The genetic locus encoding Ig is critical for vertebrate humoral immune responses and diverse antibody repertoires. Despite the critical role of buffaloes as livestock and their significance in disease transmission, the Ig loci of this species have not been thoroughly annotated. This study aimed to systematically characterize the Ig loci in buffaloes and their unique features, providing a foundation for understanding buffalo immune function.
The genomic assembly of Murrah buffalo (NDDB_SH_1) was analyzed to annotate Ig loci. Annotation criteria included functional motifs, RSS sequences, and structural features of V, D, J, and C genes. The CDR3 repertoires were constructed using genomic DNA extracted from spleen samples of five healthy buffaloes. High-throughput sequencing of multiplex PCR products enabled repertoire analysis, and MiXCR software was used for alignment and CDR3 extraction. Repertoire diversity, gene usage, and clonal frequencies were analyzed using the Immunarch R package.
The IgH locus spans approximately 667 kb on chromosome 20, containing two D-J-C clusters, 54 VH genes, 10 DH genes, 8 JH genes, and 9 CH genes. The Igκ locus, located on chromosome 12, encompasses 24 Vκ genes, 5 Jκ genes, and 1 Cκ gene, while the Igλ locus on chromosome 17 includes 71 Vλ genes, 3 Jλ genes, and 3 Cλ genes. We also conducted a detailed examination of the buffalo IgH CDR3 repertoire, revealing the presence of ultra-long CDR3 sequences, a biased usage of certain V genes, and a high-frequency usage of IgHJ1-4 genes. Furthermore, we identified a set of shared clonotypes across the samples, highlighting commonalities in the buffalo antibody repertoire.
These findings contribute to the understanding of buffalo immune function and provide insights into the evolution and diversity of ruminant immunoglobulin genes.
免疫球蛋白（Ig）是指B淋巴细胞产生的所有抗体蛋白的命名。编码Ig的基因座对于脊椎动物的体液免疫反应和多样的抗体库至关重要。尽管水牛作为家畜具有重要作用，且在疾病传播中具有重要意义，但该物种的Ig基因座尚未得到全面注释。本研究旨在系统地描述水牛的Ig基因座及其独特特征，为理解水牛的免疫功能奠定基础。
分析了穆拉水牛（NDDB_SH_1）的基因组组装以注释Ig基因座。注释标准包括功能基序、重组信号序列（RSS）以及V、D、J和C基因的结构特征。使用从五只健康水牛的脾脏样本中提取的基因组DNA构建CDR3库。多重PCR产物的高通量测序实现了库分析，并使用MiXCR软件进行比对和CDR3提取。使用Immunarch R包分析库多样性、基因使用情况和克隆频率。
IgH基因座位于20号染色体上，跨度约667 kb，包含两个D-J-C簇、54个VH基因、10个DH基因、8个JH基因和9个CH基因。位于12号染色体上的Igκ基因座包含24个Vκ基因、5个Jκ基因和1个Cκ基因，而位于17号染色体上的Igλ基因座包括71个Vλ基因、3个Jλ基因和3个Cλ基因。我们还对水牛IgH CDR3库进行了详细检查，发现存在超长CDR3序列、某些V基因的偏向使用以及IgHJ1-4基因的高频使用。此外，我们在样本中鉴定出一组共享的克隆型，突出了水牛抗体库中的共性。
这些发现有助于理解水牛的免疫功能，并为反刍动物免疫球蛋白基因的进化和多样性提供见解。
